UVB照射诱导皮肤成纤维细胞早期衰老的研究

目的 探讨UVB诱导的细胞衰老与肿瘤发生的关系。方法 噻唑蓝（MTT）法检测UVB辐射后的细胞增殖情况，筛选诱导衰老适宜的亚毒性剂量和照射次数。染色法检测衰老相关的β-半乳糖苷酶（SA β-gal）活性。RT－PCR检测衰老相关基因纤维结合素（FN）、骨结合素（ON）和平滑肌22（SM22）的表达。结果 以10 mJ/cm2的亚毒性剂量连续5次照射人成纤维细胞后，衰老的生物学特征得以明显表现：①MTT法检测显示细胞增生能力的减弱。②照射组具有SA β-gal活性的阳性细胞明显增加，照射组和对照组的阳性率分别为82.0%和33.7%（P < 0.01）。③3种衰老相关基因FN、ON和SM22的表达亦明显增强，分别约为对照组细胞的2.7、2.0、2.3倍（P < 0.05）。结论 反复亚毒性剂量UVB照射人成纤维细胞，初步建立一种UVB诱导的应激诱导的早期衰老（SIPS）模型。

Ultroviolet B exposure triggers premature senescence in human skin fibroblasts

Objective To develop a model of ultraviolet B (UVB)-induced premature senescence in human skin fibroblasts (HSF) so as to assess the relationship between stress-induced premature senescence and tumorigenesis. Methods The irradiation dose and frequency were optimized for the induction of prema-ture senescence. HSF were irradiated with UVB of 10 mJ/cm~2 once daily for 5 days, and unirradiated HSFs served as the control. After the last irradiation, cell proliferation was determined by MTT assay on day 3, 4, 5,6 and 7, SA β-Gal staining was performed to evaluate the senescence state of cells on day 3, and RT-PCR to detect the expressions of three senescene-associated genes, including fibronectin (FN), osteonectin (ON) and smooth muscle 22 (SM22) on day 3. Results After five exposures to UVB, HSF showed biological characteris-tics of senescence. As assessed by MTT assay, there was a loss of replicative potential in irradiated cells. The proportion of SA-β-gal-positive cells was 82.0% in UVB-stressed HSFs and 33.7% in the control cells (P < 0.01). The mRNA levels of FN, ON and SM22 were upregulated by 2.7, 2.0 and 2.3 folds respectively in irra-diated HSF compared with the control cells (all P < 0.05). Conclusion A stress-induced premature senes-cence model is established using HSF by repeated exposure to subcytotoxic UVB.